Promoting Homologous Recombination In Plants By Targeted cNHEJ Suppression
SUMMARY
UCLA researchers in the Department of Molecular, Cell, and Developmental Biology have developed a method to specifically suppress the highly efficient non-homologous end joining (NHEJ) pathway to boost homologous recombination efficiency in plants.
BACKGROUND
Classical non-homologous end joining (cNHEJ) and homologous recombination compete for the double-stranded DNA repair. The cNHEJ machinery recognizes DNA breaks and indiscriminately joins them, and is potentially genotoxic, whereas homologous recombination relies on the generation of 3’ overhangs, which invade homologous sister chromatids to promote error-free break repair. The choice between homologous recombination and cNHEJ depends primarily on the cell cycle stage and the nature of the break.
Recently, a cell cycle regulator of NHEJ (CYREN) was identified as a direct cell cycle-dependent inhibitor of cNHEJ that promotes error-free repair by homologous recombination during cell cycle phases when sister chromatids are present. CYREN functions by binding to the Ku heterodimer, a protein complex that is required to commence NHEJ at break sites. Thereby CYREN preferentially inhibits cNHEJ at breaks with overhangs by protecting them from Ku binding.
INNOVATION
Researchers at UCLA have developed a method to promote efficient homologous recombination directed gene targeting in plant cells. A combination of CYREN expression and creation of site specific long single stranded overhangs by the expression of CAS9-nickase will allow homologous recombination directed repair and gene targeting.
APPLICATIONS
- Accurate and efficient gene targeting and replacement in plants
- Increase the recombination efficiency for CRISPR/CAS9 technology
ADVANTAGES
Accurate and efficient gene targeting and replacement in plants.
STATE OF DEVELOPMENT
This method is ready to be tested in plant cells.
RELATED MATERIALS
N. Arnoult, A. Correia, J. Ma, A. Merlo, S. Garcia-Gomez, M. Maric, M. Tognetti, C.W. Benner, S. J. Boulton, A. Saghatelian, J. Karlseder, Regulation of DNA repair pathway choice in S and G2 phases by the NHEJ inhibitor CYREN, Nature, 2017.